Project Summary/Abstract Our long-range goal is to understand molecular pathways that govern brown fat development and function. In contrast to white fat, brown fat is a tissue that is specialized in energy expenditure. It is present in adult humans and its activity is inversely associated with human obesity. Thus, brown fat is potentially an attractive therapeutic target tissue for obesity and metabolic diseases. We are interested in how brown fat development and function is regulated by miRNAs, a class of small RNAs that do not encode protein yet control many developmental and cellular processes. We have identified a set of miRNAs that are selectively expressed in the brown fat. We found that, one of them, when expressed in the brown fat, remarkably expands the brown fat depots of mice, and as a result, the animals have less white fat mass and are resistant to diet-induced obesity. Our hypothesis is that this miRNA promotes brown fat cell differentiation by activating a cGMP- dependent signaling pathway. In the first aim, we will analyze in detail whether the expanded brown fat depots are true brown fat and are functional. We will determine whether the expanded brown fat improves metabolic parameters and enhances insulin sensitivity. We will determine whether the expanded brown fat prevents genetically predisposed obesity. In the second aim, we will perform cell culture studies to further validate that this miRNA promotes brown cell differentiation. We will examine whether the miRNA represses the expression of a negative regulator in the cGMP-dependent signaling pathway. In the third aim, we will determine whether the miRNA activates the cGMP-dependent signaling pathway. Our studies will likely provide useful information on whether mimics of this miRNA have therapeutic potentials for obesity and metabolic diseases.